Chapter 6: Replication, Maintenance, and Rearrangements of Genomic DNA
Chapter Summary
DNA REPLICATION
DNA Polymerases: Different DNA polymerases play distinct roles in DNA replication and repair in both prokaryotic and eukaryotic cells. All known DNA polymerases synthesize DNA only in the 5’ to 3’ direction by the addition of dNTPs to a preformed primer strand of DNA.
The Replication Fork: Parental strands of DNA separate and serve as templates for the synthesis of two new strands at the replication fork. One new DNA strand (the leading strand) is synthesized in a continuous manner; the other strand (the lagging strand) is formed by the joining of small fragments of DNA that are synthesized backward with respect to the overall direction of replication. DNA polymerases and various other proteins act in a coordinated manner to synthesize both leading and lagging strands of DNA.
The Fidelity of Replication: DNA polymerases increase the accuracy of replication both by selecting the correct base for insertion and by proofreading newly synthesized DNA to eliminate mismatched bases.
Origins and the Initiation of Replication: DNA replication starts at specific origins of replication, which contain binding sites for proteins that initiate the process. In higher eukaryotes, origins may be defined by chromatin structure rather than DNA sequence.
Telomeres and Telomerase: Maintaining the Ends of Chromosomes: Telomeric repeat sequences at the ends of chromosomes are maintained by the action of a reverse transcriptase (telomerase) that carries its own template RNA.
DNA REPAIR
Direct Reversal of DNA Damage: A few types of common DNA lesions, such as pyrimidine dimers and alkylated guanine residues, are repaired by direct reversal of the damage.
Excision Repair: Most types of DNA damage are repaired by excision of the damaged DNA. The resulting gap is filled by newly synthesized DNA, using the undamaged complementary strand as a template. In base-excision repair, specific types of single damaged bases are removed from the DNA molecule. In contrast, nucleotide excision repair systems recognize a wide variety of lesions that distort the structure of DNA and remove the damaged bases as part of an oligonucleotide. A third excision repair system specifically removes mismatched bases from newly synthesized DNA strands.
Translesion DNA Synthesis: Specialized DNA polymerases are capable of replicating DNA across from a site of DNA damage, although the action of these polymerases may result in a high frequency of incorporation of incorrect bases.
Recombinational Repair: Damaged DNA can be replaced by recombination with an undamaged molecule. This mechanism plays an important role in repairing damage encountered during DNA replication as well as in the repair of double strand breaks.
RECOMBINATION BETWEEN HOMOLOGOUS DNA SEQUENCES
Models of Homologous Recombination: Recombination involves the breaking and rejoining of parental DNA molecules. Alignment between homologous DNA molecules is provided by complementary base pairing. Nicked strands of parental DNA invade the other parental molecule, yielding a crossed-strand intermediate known as a Holliday junction. Recombinant molecules are then formed by cleavage and rejoining of the crossed strands.
Enzymes Involved in Homologous Recombination: The central enzyme of homologous recombination is RecA (Rad51 in eukaryotes), which catalyzes the exchange of strands between homologous DNAs. Other enzymes nick and unwind parental DNAs and resolve Holliday junctions.
DNA REARRANGEMENTS
Site-Specific Recombination: Site-specific recombination takes place between specific DNA sequences that are recognized by proteins that mediate the process. In vertebrates, site-specific recombination plays a critical role in generating immunoglobulin and T cell receptor genes during development of the immune system. Additional diversity is provided to immunoglobulin genes by somatic hypermutation and class switch recombination.
Transposition via DNA Intermediates: Most DNA transposons move throughout the genome with no requirement for specific DNA sequences at their sites of insertion. In yeasts and protozoans, however, the transposition of some DNA sequences to specific target sites results in programmed DNA rearrangements that regulate gene expression.
Transposition via RNA Intermediates: Most transposons in eukaryotic cells move by reverse transcription of RNA intermediates, similar to the replication of retroviruses. These retrotransposons include the highly repeated LINE and SINE sequences of mammalian genomes.
Gene Amplification: Gene amplification results from repeated replication of a chromosomal region. In some cases, gene amplification provides a mechanism for increasing gene expression during development. Gene amplification also frequently occurs in cancer cells, where it can result in the elevated expression of genes that contribute to uncontrolled cell proliferation.
